The present invention relates to a new and improved construction of centrifuge for separating particles suspended in a liquid, particularly although not exclusively for the separation of erythrocytes in a blood sample.
For the electronic counting of thrombocytes contained in a blood sample it is advantageous to initially remove as many erythrocytes as possible from the blood sample. The erythrocytes are more numerous and much larger than the thrombocytes so that they can disturb the measurement operation. It is known for the purpose of removing the erythrocytes to centrifuge the blood sample. The erythrocytes settle in a sediment and the remaining liquid is more suitable for counting the thrombocytes than the non-centrifuged sample.
A centrifuge is known to the art wherein the centrifuge tubes are supported at the rotor in such a fashion that their lengthwise axes intersect the rotational axis of the rotor at a constant small angle, for instance amounting to about 3.degree.. The centrifuge tubes are not pivotably arranged at the rotor. With this construction the centrifugal force acts approximately transversely with respect to the tube axis, so that in the case of centrifuge tubes containing whole blood the erythrocytes migrate along a short path (at most amounting to the diameter of the tubes) and therefore tend to correspondingly rapidly collect at the side of the tube wall facing away from the axis of rotation of the rotor. Additionally, the curvature of the tube wall favors a particularly dense collection of the erythrocytes, so that as is known there occurs an aggregation of the erythrocytes which is referred to in the art as "nummulation" or " rouleaux formation." Such formed aggregates or "rouleaux formations" possess a significantly greater sedimentation speed than the individual erythrocytes. What is disadvantageous is that the sediment, during centrifuging, deposits along the total filling height of the centrifuge tube. After stopping of the rotor the sediment migrates to the floor of the centrifuge tube. This firstly requires time and secondly causes a partial remixing or resuspension or the erythrocytes. It has been observed that sediment and residual material (remaining liquid) are not separated by a clearly defined separation surface or interface, and that the residual material, which is poor in erythrocytes, is extremely small percentually with respect to the total sample volume. The removal of a small poor in erythrocytes from this residual material is therefore extremely critical, since the currents or flow which is formed during pipetting also floats-in erythrocytes from the sediment in an uncontrolled manner. The erythrocyte concentration in the pipetted sample therefore is subjected to large fluctuations.
Another construction of centrifuge is known to the art wherein the centrifuge tubes likewise are not pivotably arranged at the rotor. The lengthwise axis of the centrifuge tubes intersects with the rotational axis of the rotor at a constant, larger angle, for instance amounting to about 30.degree.. With this arrangement the sediment already collects at the region of the tube base during centrifuging. What is disadvantageous, however, is that the erythrocytes migrate over a longer path, than in the case of an almost vertical position of the lengthwise axis of the centrifuge tube, before there is favored the rouleaux formation, so that sedimentation lasts for a longer period of time under the action of the centrifugal force. This again is disadvantageous inasmuch as loading of the cells by the action of the centrifugal force causes a morphological change of the cells which is progressive in the static sense. Additionally, also disadvantageous is the fact that during centrifuging the separation surface or interface which forms between the sediment and the remaining or residual material extends approximately vertically, and therefore, at an inclination to the lengthwise axis of the centrifuge tube. If the centrifuge tube, following stoppage of the rotor, remains for a while in situ, then there is formed a horizontal separation surface, which, however, likewise is inclined with respect to the lengthwise axis of the centrifuge tube. If the centrifuge tube is removed from the centrifuge immediately upon standstill of the rotor or later, then it is placed in a stand which normally is provided for a vertical tube axis, and there is again formed a horizontal separation surface, i.e., extending transversely with respect to the tube axis. As should be apparent there occurs at least once a remixing effect, which, while less prounouced than with the first described centrifuge construction, still the previously mentioned drawbacks essentially are also present here.
Additionally, a centrifuge has been disclosed to the art wherein the centrifuge tubes are pivotably mounted and can shift, under the action of the centrifugal force, from a vertical position into an almost horizontal position. With this arrangement the centrifugal force acts, as desired, in the direction of the tube axis and the tube floor or base. What is disadvantageous however is that the erythrocytes migrate over a longer path, which can amount to the total filling height of the centrifuge tube. This counteracts or completely elminates the rapid rouleaux formation if the sample consists of diluted blood, since the rouleaux formation only arises at those locations where the dilution of the erythrocytes does not exceed a ratio of about 1:5. Furthermore, what is additionally disadvantageous is that the displacement flow, caused by migration of the cells, counteracts sedimentation. In order to obtain the necessary degree of separation there is thus required a long centrifuging time, which, in turn, again increases the morphological alteration of the cells.
There is attained a proper separation of the particles from the liquid and there is extensively avoided the remixing effect if the centrifuge tubes, during standstill of the rotor, reposition themselves into such a rest position under the influence of the force of gravity that their lengthwise axes are essentially vertically oriented, and further, if the centrifuge tubes and their holders, during the centrifuging action, pivot-out until reaching a stop or impact member, under the combined effect of the force of gravity and the centrifugal force, so that the lengthwise axes of the centrifuge tubes assume a predetermined inclined position. During run-up and run-down of the rotor the lengthwise axes of the centrifuge tubes, corresponding to the momentarily prevailing centrifugal force, continuously shift from the vertical or inclined position into the inclined or vertical position, as the case may be. Thus, during centrifuging the sediment is collected at the region of the floor or base of each tube. Morever, the path through which the particles must move until rouleaux formation is not as long as in the case of centrifuging with almost horizontal disposition of the lengthwise axes of the centrifuge tubes. During run-down of the rotor the separation surface between the sediment and the liquid, which is vertical during centrifuging and thus inclined with respect to the lengthwise axis of the centrifuge tubes, slowly and continuously shifts into the horizontal position which at the same time is located perpendicular to the lengthwise axis of each centrifuge tube. The remixing effect is extensively avoided by virtue of this slow and continuous positional change of the centrifuge tubes, and this is even more so by virtue of the fact that the centrifugal force, after the point in time where the holders no longer contact the stops, always acts in the direction of the lengthwise axes of the centrifuge tubes. Upon stoppage of the rotor the centrifuge tubes are oriented so that each lengthwise axis is vertically dispositioned. There are thus combined the advantages of the individually known centrifuge methods, without having to take into account their drawbacks. The end result of such is that, on the one hand, there is obtained a shorter centrifuging time, and thus, a lesser morphological change of the particles (if such for instance are erythrocytes), and, on the other hand, there is realized an optimum suppression of the remixing between the sediment and the remaining liquid.
It is however necessary to be able to adjust the inclined position of the centrifuge tubes between almost 0.degree. and 90.degree. in order to optimumly accommodate the momentary requirements of the samples to be centrifuged (in accordance with the blood dilution, the tube diameter and so forth). From U.S. Pat. Nos. 2,739,759, 3,722,789 and 3,951,334 there have become known to the art centrifuges wherein there are adjustably arranged at the rotor screws serving as stops for limiting the outward pivoting or rocking of the centrifuge tubes and their holders. These known stops must be however individually adjusted, so that it is difficult to avoid different settings of the stops and the corresponding imbalance at the rotor. Also, such type stops can tend to alter their setting during centrifuging, and finally, it is cumbersome to change the setting of the stops for accommodating different requirements and to again properly reset the stops to a previous setting after a change has been made. Furthermore, from the aforementioned U.S. Pat. No. 2,739,759 it is known to construct the stops in each case as contact locations of a disk-shaped body equipped with slots and the pivoted-out centrifuge tubes. During the pivoting-out movement each respective centrifuge tube is radially guided in one of the slots. With this design the stops or impact members however are not adjustable.